Thrombogenesis with continuous blood flow in the inferior vena cava

Abstract

SummarySeveral rodent models have been used to study deep venous thrombosis (DVT). However, a model that generates consistent venous thrombi in the presence of continuous blood flow, to evaluate therapeutic agents for DVT, is not available. Mice used in the present study were wild-type C57BL/6 (WT), plasminogen activator inhibitor-1 (PAI-1) knock out (KO) and Delta Cytoplasmic Tail (ΔCT). An electrolytic inferior vena cava (IVC) model (EIM) was used. A 25G stainless-steel needle, attached to a silver coated copper wire electrode (anode), was inserted into the exposed caudal IVC. Another electrode (cathode) was placed subcutaneously. A current of 250 μAmps over 15 minutes was applied. Ultrasound imaging was used to demonstrate the persence of IVC blood flow. Analyses included measurement of plasma soluble P-selectin (sP-Sel), thrombus weight (TW), vein wall morphometrics, P-selectin and Von Willebrand factor (vWF) staining, transmission electron microscopy (TEM), scanning electron microscopy (SEM); and the effect of enoxaparin on TW was evaluated. A current of 250 μAmps over 15 minutes consistently promoted thrombus formation in the IVC. Plasma sP-Sel was decreased in PAI-1 KO and increased in ΔCT vs. WT (WT/PAI-1: p=0.003, WT/ΔCT: p=0.0002). Endothelial activation was demonstrated by SEM, TEM, P-selection and vWF immunohistochemistry and confirmed by inflammatory cell counts. Ultrasound imaging demonstrated thrombus formation in the presence of blood flow. Enoxaparin significantly reduced the thrombus size by 61% in this model. This EIM closely mimics clinical venous disease and can be used to study endothelial cell activation, leukocyte migration, thrombogenesis and therapeutic applications in the presence of blood flow.

The electrolytic inferior vena cava model (EIM) to study thrombogenesis and thrombus resolution with continuous blood flow in the mouse

“…The thrombus shape confirms thrombus formation in the presence of blood flow (14, 15). Initially, chronic time points were not evaluated (14, 15). Here, we present a follow-up study on the EIM in order to determine whether the IVC remains with blood flow from acute through chronic time points, allowing for the study of not only the thrombus formation but also of thrombus resolution.…”

“…Male (n=195) wild-type (WT) C57BL/6 mice (Charles River Laboratories, Wilmington, MA, USA), weighing 20 to 25 grams, were anesthetised with isoflurane 2%, placed in dorsal recumbency and the IVC was approached directly via a midline laparotomy utilizing aseptic technique (14, 15). Venous side branches were ligated using 7-0 Prolene suture (Ethicon, Inc, Somerville, NJ), while back branches remained patent.…”

“…Thus, we developed the electrolytic IVC model (EIM), a flow model that produces a non-occlusive and consistent IVC thrombus in the presence of preserved blood flow (14, 15). Using the EIM, we have demonstrated preserved IVC endothelium are activated, causing P-selectin expression, which in turn leads to white blood cell attachment and migration into the vein wall (▶Figure 1) with red blood cells and fibrin deposition contributing to the growing thrombus (14). The thrombus shape confirms thrombus formation in the presence of blood flow (14, 15).…”

“…Using the EIM, we have demonstrated preserved IVC endothelium are activated, causing P-selectin expression, which in turn leads to white blood cell attachment and migration into the vein wall (▶Figure 1) with red blood cells and fibrin deposition contributing to the growing thrombus (14). The thrombus shape confirms thrombus formation in the presence of blood flow (14, 15). Initially, chronic time points were not evaluated (14, 15).…”

Animal Models of Thrombosis From Zebrafish to Nonhuman Primates

“…102 Free-radical injury applied to the outer surface or intraluminally to the endothelial surface is a general approach for thrombus induction in mouse veins, using either ferric chloride 31,67 or electrolytic injury mechanisms. 56 The latter have shown more consistency when applied to the femoral vein 68,75 or IVC 103 to generate both acute and more chronic thrombi. Other models of acute thrombosis have included pinch injury, 69 intraluminal insertion of collagen-dominated surfaces, 61 and microsurgical anastomosis.…”